The host innate immune response is triggered within hours of virus infection. As a whole, its function is to limit virus replication at local sites of infection and to orchestrate development of the adaptive immune response. Viruses are typically recognized by cellular pattern recognition receptors (PRRs), including toll-like receptors (TLRs) and the retinoic acid inducible gene (RIG)-like RNA helicases (RLHs). Ligation of these PRRs, often by viral nucleic acids, culminates in the activation of multiple transcription factors that cooperate in driving expression of cytokines and chemokines characteristic of the innate response. Nuclear factor-kappa B (NF-kappaB) and interferon (IFN) regulatory factors (IRFs) are particularly important transcription factors, responsible for induction of type I IFN (IFNalpha/beta), tumor necrosis factor alpha (TNFalpha) and other mediators of inflammation. IFNalpha/beta is central to the anti-viral response as it initiates its own transcriptional program resulting in expression of IFN-stimulated genes (ISGs) via the Janus kinase-signal transducer and activation of transcription (JAK-STAT) pathway. ISG expression influences many cellular processes including RNA processing, protein stability and cell viability that can directly affect virus replication. ISG expression in cells of the immune system such as dendritic cells (DCs) and macrophages is critical for antigen presentation and T- and B-cell activation, thus affecting the quality of the adaptive immune response and eventual virus clearance. To facilitate dissemination, pathogenic viruses have evolved mechanisms to suppress host innate immunity by antagonizing these signal transduction pathways. Hence, understanding the specific pathways by which viruses activate and evade innate immune responses is essential for understanding viral pathogenesis as well as for development of effective vaccines. To examine virus-host interactions that affect innate immunity, our laboratory utilizes flaviviruses as the primary model of infection. Flaviviruses have an essentially global distribution and represent a tremendous disease burden to humans, causing millions of infections annually. The success of flaviviruses as human pathogens is associated with the fact that they are arthropod-borne, transmitted by mosquitoes or ticks. Significant members of this group include dengue virus (DENV) and yellow fever virus (YFV) that cause hemorrhagic fevers, as well as Japanese encephalitis virus (JEV), West Nile virus (WNV) and tick-borne encephalitis virus (TBEV) that cause severe encephalitides. These viruses are listed as NIAID category A, B and C pathogens for research into their basic biology and host response. The flavivirus single-stranded RNA genome is translated as one open reading frame; the resulting polyprotein is cleaved into at least ten proteins that include three structural (capsid C, membrane M, derived from the precursor preM and envelope E), and seven nonstructural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5). Virus replication proceeds in association with modified membranes derived from the endoplasmic reticulum of host cells. NS5 is the largest and most conserved of the flavivirus proteins containing approximately 900 amino acids. It encodes a methyltransferase (MTase) and RNA-dependent RNA polymerase (RdRP) and associates with NS3 (the viral protease) to form the functional unit of the viral replication complex. Despite the widespread and often severe infections caused by these pathogens, vaccines exist for only a few (YFV, JEV and TBEV) and no therapeutic exists to treat clinical infection caused by any flavivirus. Type I IFNs are essential to recovery from flavivirus infection and have been used clinically as potential therapeutics, albeit with limit success. This may be due to the observation that all flaviviruses examined to date antagonize IFN-dependent responses by suppressing JAK-STAT signal transduction. We identified NS5 as the major IFN antagonist encoded by flaviviruses, originally using Langat virus (LGTV; a member of the TBEV complex of flaviviruses) and most recently using WNV. Although other NS proteins contribute to suppression of JAK-STAT signaling, studies by our laboratory and others suggest that NS5 is the most potent of the IFN antagonist proteins encoded by all vector-borne flaviviruses examined thus far. Hence, determining the mechanism(s) by which NS5 impedes signaling is essential to understand flavivirus pathogenesis and may lead to new therapeutic targets. Furthermore, it is important to understand the mechanisms underlying the anti-viral effects of IFN by identifying the function of ISGs with anti-viral activity. Finally, it is essential to translate these findings to immunologically relevant cell types and animal models to understand the roles of induction and evasion of innate immunity in development of the adaptive immune response and in virus pathogenesis. Achieving these goals will significantly improve our understanding of how viruses cause disease and identify therapeutic targets for intervention.